Summary This application explores the functions of a DNA repair factor as a new element affecting evolution and adaptation of bacteria, including those that cause infections. This is of scientific interest because it involves the discovery of molecular pathways that affect bacterial cell physiology and evolution in conditions of stress. Thus, the knowledge generated by this application has the potential to uncover novel targets for the treatment of pernicious bacterial infections. The hypothesis under study is that the Mfd factor, previously studied in the context of high-fidelity DNA repair of transcribed regions, mitigates the toxic effects associated with oxidative stress by mediating error-prone DNA repair of DNA lesions and activating expression of genes that prevent cell death. This hypothesis will be tested using genetic, biochemistry, and systems biology approaches. In Aim 1, we will test the concept that Mfd-dependent mutagenesis operates at genes that are highly transcribed in cells experiencing oxidative stress. This concept will be examined innovatively by measuring mutagenesis at the genomic level and in the absence of selection. Aim 2 will elucidate whether Mfd is important for activating expression of factors that protect against protein damage caused by oxidative stress independently of its previously associated DNA repair functions. In conclusion, the experiments in this application are straight-forward and self- contained and seek to determine new functions of the Mfd factor. Mfd is well-conserved in bacterial pathogens and so the research proposed can potentially uncover a novel therapeutic target.